Stroke is the third leading cause of death and the leading cause of disability in the United States. For patients with ischemic stroke, the thrombolytic drug recombinant tissue plasminogen activator (rt-PA) is the only FDAapproved treatment. Systemic or peripheral administration of rt-PA has been used in the treatment of stroke with limited success as it is difficult to control delivery ofrt-PA to the target clot through a peripheral injection. Direct administration of rt-PA into the clot through a catheter shows promise, but to date has been limited by technical demands and availability of expertise in its administration. In addition, rt-PA has a significant risk to induce intracerebral hemorrhage. Therefore, to improve the efficacy of rt-PA-induced clot lysis and to reduce the administered rt-PA dose, we have investigated the synergistic effect of rt-PA and 120-KHz ultrasound on thrombolysis in an in vitro porcine clot model. These preliminary data strongly support the central hypothesis of our proposal that ultrasound enhances thrombolysis, primarily via mechanical mechanisms. To test this hypothesis we propose to investigate three Specific Aims: In Aim #1, we will expand the knowledge base of mechanisms of ultrasound-mediated thrombolysis.in a series of in vitro experiments with a porcine clot model. Using a 30-MHz active cavitation detector (or 15-MHz passive...Constantin?), we will determine the relationship between cavitational activity and accelerated thrombolysis from exposure to pulsed ultrasound. Thermocouple measurements of the clot temperature during the quiescent period of pulsed ultrasound will elucidate the contribution of thermal effects to ultrasound-enhanced thrombolysis. As a novel approach in Aim #2, we will investigate the potential of echogenic liposomes (ELIP), or submicron-sized phospholipid bilayer vesicles enclosing gas and fluid developed by McPherson and MacDonald at Northwestern University, to deliver rt-PA near the intravascular clot causing the ischemic stroke. Furthermore, ELIP can be targeted to clot by conjugating anti-fibrin antibodies to the liposome surface. Diagnostic ultrasound (low MI) will be used in vitro to identify the t-PA-loaded liposomes and to assess the targeting efficiency. The threshold of liposome destruction and drug delivery will be determined using a clinical ultrasound scanner (ATL HDI 5000). Lastly, in Aim #3, we will conduct in vivo studies in porcine hemorrhagic and ischemic stroke models to demonstrate the efficacy of ultrasound-enhanced thrombolysis and to clarify the potential risks for therapeutic ultrasound on brain tissue and in the presence of an echo contrast agent. We expect that successful completion of these studies will contribute significantly to our long-term goal to develop an ultrasound-assisted thrombolysis system that delivers and enhances thrombolytic therapy in the cerebral vasculature and rapidly restores perfusion after ischemic stroke.